This investigation of the mechanism and structure of the (Na ion plus K ion)-ATPase is proceeding along three lines: (a) determination of the transient kinetics of phosphorylation and dephosphorylation of the enzyme; (b) ligand-binding studies; and (c) subunit structure studies. The transient kinetic studies have shown that the mechanism of hydrolysis proceeds through the phosphorylated intermediate, which in turn produces a conformational transition. The ligand binding studies have shown that the stoichiometry of ouabain-binding sites relative to phosphorylation sites in the Electrophorus electric organ preparation differs from that of the mammalian enzymes and we are investigating the basis for this difference. The subunit studies have developed higher resolution techniques for separating the peptide chains and shown that the purified alpha- and beta-chains are heterogenous by these techniques. We now seek to determine whether this is an indication of complex structure of isozymes.